Protease inhibitors are key components in the chemotherapy of HIV/AIDS. Unfortunately, the long term efficacy of protease inhibitors is severely compromised by the appearance of drug-resistant mutations that lower their potency to inadequate levels for effective inhibition and viral suppression. The onset of drug resistance is often accelerated by issues of patient compliance, often aggravated by severe side effects. In addition, the viral subtypes prevalent in Africa, where the vast majority of HIV infections take place, are not the same as the one responsible for the infections in America and Europe. Complicating things even further, a different HIV virus, HIV-2, although less prevalent than HIV-1, is also able to cause AIDS. It is evident, that the development of new protease inhibitors with high potency, effectiveness against different subtypes, low susceptibility to mutations and minimal side effects still remains an urgent goal. The main goal of this project is to develop precise thermodynamic and structural guidelines to develop such inhibitors. The specific goals of this project are: - Identification of thermodynamic and structural determinants of extremely high affinity. - Identification of thermodynamic and structural determinants that confer protease inhibitors low susceptibility to mutations and efficacy against different viral subtypes, including HIV-2. - Identification of thermodynamic and structural determinants that lower the affinity of protease inhibitors to unwanted targets and hence improve selectivity and reduce side effects. The goals will be achieved by a combination of experimental thermodynamic measurements (high sensitivity isothermal titration calorimetry and high sensitivity differential scanning calorimetry), structure determination (x-ray crystallography) and structure-based thermodynamic analysis.